Analysis of the physical chemistry of linear DNA containing replication loops has demonstrated that the loops are remarkably unstable and are extruded as double stranded DNA at a significant rate even at temperatures 50 degrees and more below the Tm. The rate limiting step in this extrusion is the rate of end melt and/or nucleation of the nascent strands, not the rate of branch migration. This unusual feature provides a general method for the isolation of origins of DNA replication. In other experiments we have demonstrated that SV40 is not a potent mutagen and that is weak mutagenic activity is probably unrelated to its ability to transform.